The Agouti Related Protein (AGRP) is upregulated in obese and diabetic mice and stimulates hyperphagia and the development of obesity when overexpressed in transgenic mice. The human ortholog, hAGRP, has been isolated and has similar molecular and physiological properties, expressing one transcript in the arcuate nucleus and one in the periphery. We recently determined the gene structure and identified the minimal promoter of the human AGRP gene. Multiple binding sites for transcription factors were identified including putative binding sites for the STAT transactivators that may potentially mediate leptin's action in the hypothalamus. The 5' non-coding exon had significant promoter activity in a periphery cell line only, suggesting its role in the expression of the periphery-specific transcript. We also identified a polymorphism in the promoter that had significant impact on promoter activity and affinity to bind transcription factors, as tested in hypothalamus- and periphery-derived cell lines. The genotype with the high promoter activity was significantly associated with obesity and Type 2 Diabetes Mellitus (T2DM) in two different cohorts. Moreover, a polymorphism in the coding region of the gene was significantly associated with low body mass index and low abdominal adiposity but in older people only, suggesting a role for hAGRP in preventing late on set obesity. We hypothesize that increased amounts of hAGRP in the hypothalamus and the periphery, as determined by its promoter, will result in hyperphagia and the development of obesity. We therefore propose to completely characterize the promoter of hAGRP and identify the transcription factors and sequence motifs that regulate expression of the gene (Specific Aim 1). We will also determine the impact of hormones/fuel substrates on promoter activity and endogenous expression of AGRP, and identify the hormone/substrate response elements in the promoter of the gene (Specific Aim 2). Furthermore, we will investigate the association of hAGRP polymorphisms with obesity and T2DM, while the systemic levels of AGRP will be correlated with promoter polymorphisms as well as with the systemic levels of hormones/fuel substrates (Specific Aim 3). Given that hAGRP is a potent appetite effector, it is important that we identify the regulatory mechanisms for its expression and determine its role in food intake and the development of human obesity.